Cholesterol is an important constituent of cells and is also a clinically important constituent since an excessive level of cholesterol causes transformation of macrophages into foam cells after uptake thereof by macrophages in the subendothelial space and then causes development of a primary lesion of arteriosclerosis. Low density lipoprotein (LDL) play a major role in cholesterol transport in the blood and are risk factors for arteriosclerosis. It is known that small, dense LDL, which are particularly small in particle size among LDL and higher in density compared with standard LDL, is more several fold atherogenic than normal LDL. Increase of small, dense LDL is one of the major risk factors for arteriosclerosis. It is clinically very important to perform a fractional measurement for such small, dense LDL.
Examples of conventional methods for measurement of small, dense LDL include an ultracentrifugation method, an electrophoresis method, and a method using high performance liquid chromatography. These methods are not convenient since they require expensive facilities and much time for measurement.
An example of a method for measuring small, dense LDL using an autoanalyzer is a method (see JP Patent Publication (Kokai) No. 2003-28882 A) that involves suspending or dissolving small particle LDL with the use of differences in ionic strength and then conducting measurement on the small particle LDL with the use of differences in absorbance. However, differences in absorbance are measured based on turbidity according to such method. Hence, cholesterol in small, dense LDL cannot be measured and thus specificity and accuracy are insufficient.
Also, a method (International Patent Publication WO2004/053500) that involves measuring cholesterol or triglyceride in small, dense LDL through the use of a combination of a separating agent comprising polyanions and a divalent cation and a reagent adaptable for an autoanalyzer is known. This method is capable of measuring lipid components in small, dense LDL more conveniently than an ultracentrifugation method or an electrophoresis method. Furthermore, the method is excellent in specificity and accuracy. However, the method requires pretreatment of specimens and a procedure for separating LDL into small, dense LDL and LDL other than such LDL.